Pathogens employ many strategies to evade immune recognition. Correspondingly, the immune system has evolved antigen receptors and sensors for molecular patterns to recognize microbial invasion. Despite these efforts, microbial detection may be inadequate and severe infection results. IL-1? release from DCs and macrophages occurs when cytosolic microbial products or cellular stress are detected by specialized sensors capable of assembling into inflammasomes. An orchestrated series of events occurs since pro-IL-1? must be cleaved to its active form and lacking a signal peptide, must be released directly from the cytosol. Inflammasome assembly results in caspase-1 activation with subsequent cleavage of pro-IL-1?, Gasdermin-D and pyroptotic cell lysis that releases bioactive IL-1?. However, many microbes evade inflammasome activation and IL-1? release. A special group of T cells, invariant (i) NKT cells sits at the interface between innate and adaptive immunity. They have a semi-invariant T cell receptor that recognizes microbial or TLR-activated self-lipid antigens presented by CD1d on DCs. iNKT cells are ?cellular adjuvants? because they recognize early infection, secrete high levels of cytokines, and trans-activate other leukocytes to amplify stronger and more rapid immune responses. Here, we define an axis in which iNKT cells drive IL-1? release from DCs when inflammasome activation fails. The mechanism involves Fas-FasL engagement that instead of apoptotic cell death, results in a special state of cellular activation referred to as ?hyperactivation? of DCs, where sustained bioactive IL-1? release occurs from living cells. In Aim 1, we determine how IL-1? is released from TLR-primed DCs following Fas ligation. We hypothesize that IL-1? may escape via cell membrane pores resulting from activation of gasdermins (besides gasdermin-D). We will use CRISPR/Cas9 and RNAi methods to delete or silence genes encoding various caspase and gasdermins to determine those required for pro-IL-1? cleavage, pore formation and release. In Aim 2, we determine how DCs escape apoptotic cell death after Fas ligation. Based on preliminary data, we hypothesize that this occurs by rapid proteasomal degradation of active caspase or gasdermin effectors. We will determine the effects of proteasome inhibition and which caspases and related cell death mediators become predominantly K11 and K48-ubiquitin-linked to learn how cell death is avoided. In Aim 3, we test commensal (B. fragilis) and pathogenic (C. trachomatis and S. typhimurium) live bacterial infection of DCs in vitro in the presence or absence of iNKT cells. Based on preliminary data that several of these infections result in IL-1? release only if iNKT cells are present, we will determine if the mechanism responsible is CD1d dependent, Fas mediated, and if bioactive IL-1? is released through membrane pores similar to the mechanism established in the reductionist models in Aims 1 and 2.